Spiral NeuroString: High-Density Soft Bioelectronic Fibers for Multimodal Sensing and Stimulation.

Bioelectronic fibers hold promise for both research and clinical applications due to their compactness, ease of implantation, and ability to incorporate various functionalities such as sensing and stimulation. However, existing devices suffer from bulkiness, rigidity, limited functionality, and low density of active components. These limitations stem from the difficulty to incorporate many components on one-dimensional (1D) fiber devices due to the incompatibility of conventional microfabrication methods (e.g., photolithography) with curved, thin and long fiber structures. Herein, we introduce a fabrication approach, ‶spiral transformation″, to convert two-dimensional (2D) films containing microfabricated devices into 1D soft fibers. This approach allows for the creation of high density multimodal soft bioelectronic fibers, termed Spiral NeuroString (S-NeuroString), while enabling precise control over the longitudinal, angular, and radial positioning and distribution of the functional components. We show the utility of S-NeuroString for motility mapping, serotonin sensing, and tissue stimulation within the dynamic and soft gastrointestinal (GI) system, as well as for single-unit recordings in the brain. The described bioelectronic fibers hold great promises for next-generation multifunctional implantable electronics.

Stem cell activation during distraction enterogenesis in the murine colon.

PURPOSE: Short bowel syndrome (SBS) is a devastating disease. We have proposed spring-mediated distraction enterogenesis for intestinal lengthening. Colonic lengthening is a potential treatment option for SBS to enhance fluid absorption capacity. We hypothesized that intraluminal spring-mediated colonic lengthening is associated with stem cell proliferation. METHODS: C57BL/6 mice underwent placement of a gelatin-encapsulated compressed or uncompressed nitinol spring in a cecal segment. Animals were given clear liquid diet until postoperative day (POD) 7, followed by regular diet until POD 14. Cecal lengths were measured at euthanasia, and tissue was formalin fixed for histological processing. For Lgr5-GFP mice, immunohistochemistry against GFP was performed to localize Lgr5+ cells within crypts. RESULTS: Significant cecal lengthening with compressed springs and shortening with uncompressed springs were observed on POD 7 and 14. Mucosa of the compressed spring group was significantly thicker on POD 14. The density of Lgr5+ cells within the crypts in the compressed spring groups was higher than that in the uncompressed spring groups on both POD 7 and 14. CONCLUSION: Expandable springs can be used to lengthen the colon in the mouse model. Colonic lengthening was associated with gradual mucosal thickening and correlated with an increased density of stem cells within the crypts.

Long-term safety of intraluminal spring-mediated bowel lengthening.

PURPOSE: The purpose of the study is to examine the long-term safety of an endoluminal bowel lengthening device prior to its use in the first human trial. In addition, device performance and natural passage will be evaluated. METHODS: Endoluminal lengthening springs were surgically placed into the jejunum of Yucatan minipigs using the Eclipse XL1 device. A matching internal control segment of jejunum was marked at the time of operation. Weekly weights and fluoroscopic studies were obtained to evaluate spring deployment and position until devices passed. Animals were euthanized at 28, 60, 90, and 180 days. At necropsy, length measurements were recorded, and histopathologic analysis was performed. RESULTS: There were no bowel obstructions or overt perforations attributable to the device. All surviving animals gained weight and were clinically thriving. All devices passed out of the rectum by 180 days. Bowel lengthening was seen in all experimental segments, and minimal fibrosis was observed by 180 days. CONCLUSION: Jejunal lengthening persisted after device had passed through the intestinal tract after 180 days. Early histopathologic changes of the jejunum during distraction enterogenesis normalized over time.

The effect of spring diameter on porcine ileal distraction enterogenesis.

PURPOSE: Spring-mediated distraction enterogenesis has proven to be successful for intestinal lengthening. We aimed to evaluate the effect of spring diameter mismatch on intestinal adaptation. METHODS: Juvenile mini-Yucatan pigs underwent placement of compressed nitinol springs with diameter of 10, 11, or 12 mm into the ileal lumen. Pigs were euthanized on postoperative day 7. The lengths, histology, total area of blood vessels, and enteric ganglia were evaluated. RESULTS: All spring groups exhibited significant ileal lengthening. Across the different diameters, spring-expanded segments were similar in terms of ileal lengthening, crypt height, muscular thickness, blood vessels, and enteric ganglia area. CONCLUSION: Spring-mediated distraction enterogenesis is successful in the porcine ileum. A smaller diameter spring is as effective as a larger diameter spring in lengthening the ileum. Springs of varying diameters result in comparable structural changes in the ileum.

Internal plication for spring confinement to lengthen intestine in a porcine model.

BACKGROUND: Short bowel syndrome and its resultant nutritional deficiencies are the most common cause of intestinal failure. Significant intestinal lengthening using intraluminal springs is feasible in porcine models using an external plication technique. We hypothesize that an internal plication technique will yield significant intestinal lengthening, which may lead to future endoscopic spring placement. METHODS: Uncompressed springs measuring 7.5 cm with a diameter of 1.0 cm were compressed to 2.0 cm. A gelatin-encapsulated compressed nitinol spring was inserted into the jejunal lumen of juvenile pigs and held in place with endoluminal sutures just proximal and distal to the spring-containing segment. A control segment distal to the spring was marked. Pigs were euthanized on postoperative day 7. Spring and control segments were collected for analyses. RESULTS: There was an average lengthening by 72% of the spring segment compared to the control segment. Two out of 7 springs stayed within both sets of plications and doubled in length. Histology showed normal mucosal integrity of the spring segment and plicated areas with similar muscular thickness but increased crypt depth and villus length compared to the control segment. CONCLUSION: Internal plication resulted in significant bowel lengthening. Five springs had slipped through proximal, distal or both sets of plications, resulting in less lengthening than those that remained fixed. A more consistent methodology for endoluminal suturing is needed to produce more lengthening.

Generation of Porcine Ileum Through Spring-Mediated Mechanical Distraction.

INTRODUCTION: Short bowel syndrome is a devastating gastrointestinal disorder in which decreased bowel length results in inadequate absorption causing nutritional deficiencies. Current treatment options are accompanied by significant morbidity. We have proposed spring-mediated distraction enterogenesis as a method to lengthen bowel with success seen in porcine jejunum. We hypothesize that spring-mediated distraction enterogenesis can be demonstrated in porcine ileum with preservation of ileal structure and function. MATERIALS AND METHODS: Laparotomy was performed on juvenile female mini-Yucatan pigs and a gelatin-encapsulated compressed nitinol spring was inserted into the ileal lumen and affixed proximally and distally. A control segment distal to the spring segment was marked with sutures. Postoperatively, pigs were placed on a liquid diet and euthanized on postoperative day 7. Spring and control segments were measured and processed for immunohistochemistry to evaluate for the presence of vitamin B12-intrinsic factor cotransporter, chromogranin A-producing cells, and 5-HT producing cells. RESULTS: All seven pigs survived to postoperative day 7 with no adverse effects. On average, pigs gained 84.3 ± 66.4 g/d. Spring segments lengthened 1.5 ± 0.7 cm with a relative lengthening by 128% ± 56%, which was statistically significant when compared to control (P < 0.01). The average density of chromogranin-A cells in control compared to spring segments was not significantly changed (2.9 ± 1.1 cells/mm versus 3.2 ± 1.2 cells/mm, P = 0.17). Both vitamin B12-intrinsic factor cotransporter and 5-HT producing cells were present in both control and lengthened ileum. CONCLUSIONS: Intraluminal nitinol springs significantly lengthened porcine ileum. The increase in density of enteroendocrine cells may indicate enhanced endocrine function of the lengthened ileum.

A tissue-like neurotransmitter sensor for the brain and gut.

Neurotransmitters play essential roles in regulating neural circuit dynamics both in the central nervous system as well as at the peripheral, including the gastrointestinal tract1-3. Their real-time monitoring will offer critical information for understanding neural function and diagnosing disease1-3. However, bioelectronic tools to monitor the dynamics of neurotransmitters in vivo, especially in the enteric nervous systems, are underdeveloped. This is mainly owing to the limited availability of biosensing tools that are capable of examining soft, complex and actively moving organs. Here we introduce a tissue-mimicking, stretchable, neurochemical biological interface termed NeuroString, which is prepared by laser patterning of a metal-complexed polyimide into an interconnected graphene/nanoparticle network embedded in an elastomer. NeuroString sensors allow chronic in vivo real-time, multichannel and multiplexed monoamine sensing in the brain of behaving mouse, as well as measuring serotonin dynamics in the gut without undesired stimulations and perturbing peristaltic movements. The described elastic and conformable biosensing interface has broad potential for studying the impact of neurotransmitters on gut microbes, brain-gut communication and may ultimately be extended to biomolecular sensing in other soft organs across the body.

Gastrointestinal Myoelectric Measurements via Simultaneous External and Internal Electrodes in Pigs.

INTRODUCTION: Currently, there is no accurate noninvasive measurement system to diagnose gastrointestinal (GI) motility disorders. Wireless skin patches have been introduced to provide an accurate noninvasive measurement of GI myoelectric activity which is essential for developing neuro-stimulation devices to treat GI motility disorders. The aim of this study is to compare the external and internal electrical signal measurements in ambulatory pigs. METHODS: Yucatan pigs underwent placement of internal electrodes on the stomach, small intestine, and colon. Wires were brought through the abdominal wall. Signals were collected by a wireless receptor. Four external patches were placed on the abdominal skin to record the signals simultaneously. Pigs were kept for 6 d while the sensors were continuously recording the data from both systems. RESULTS: Internal sensors detected rich signals from each organ. The stomach had a dominant frequency that ranged from 4 to 4.5 cpm, with occasional higher frequencies at 2, 3 and 4 times that. Small intestine signals had their primary energy in the 12-15 cpm range. Colon signals primarily displayed a dominant broad peak in the 4-6 cpm region. External skin patches detected a substantial fraction of the activities measured by the internal electrodes. A clear congruence in the frequency spectrum was observed between the internal and external readings. CONCLUSIONS: Internally measured myoelectrical signals confirmed different patterns of rhythmic activity of the stomach, small intestine, and colon. Skin patches provided GI myoelectric measurement with a range of frequencies that could be useful in the diagnosis and treatment of motility disorders.

Distraction enterogenesis in the murine colon.

BACKGROUND/PURPOSE: Distraction enterogenesis with intraluminal spring technology has been successfully used to lengthen segments of murine small intestine. We hypothesized that biocompatible springs could also be used to lengthen murine large intestine. METHODS: Age and weight matched C57BL/6 mice underwent surgical insertion of nitinol spring-loaded capsules into the cecum. Segment lengths were measured at initial spring placement and at euthanasia after 7 and 14 days. Histologic adaptations were evaluated at scarification. RESULTS: Cecal segments loaded with compressed springs lengthened an average of 150%, which was significantly longer than control segments loaded with either empty capsules or uncompressed springs. Muscularis layers tended to be thicker in the compressed spring groups compared to control groups. CONCLUSIONS: Insertion of a compressed nitinol spring into the cecum results in significant colonic lengthening in a mouse model. The ability to increase cecum length serves as proof of concept that distraction enterogenesis technology may be feasibly applied to large intestinal models. The use of distraction enterogenesis technology shows promise for application to clinical models in the treatment of pediatric intestinal disease.

Mechanical lengthening of porcine small intestine with decreased forces.

INTRODUCTION: short bowel syndrome is marked by inadequate intestinal surface area to absorb nutrients. Current treatments are focused on medical management and surgical reconfiguration of the dilated intestine. We propose the use of spring-mediated distraction enterogenesis as a novel intervention to increase intestinal length. Given our previous success lengthening intestinal segments using springs with spring constant ~7 N/m that exerts 0.46 N or higher, we sought to determine the minimal force needed to lengthen porcine small intestinal segments, and to explore effects on intestine over time. METHODS: Juvenile Yucatan pigs underwent laparotomy with enterotomy to introduce nitinol springs intraluminally (n = 21 springs). Bowel segments (control, spring-distracted) were retrieved on post-operative day (POD) 7 and 14, and lengths measured. Thickness of cross-sectional intestinal layers were measured using H&E, and submucosal collagen fiber orientation measured using trichrome stained sections. RESULTS: all pigs survived to POD7 and 14. Spring constants of at least 2 N/m exerting a minimum force of 0.10 N significantly lengthened intestinal segments (p <0.0001). The stronger the spring force, the greater the induced thickness of various intestinal layers at POD7 and 14. Collagen fiber orientation was also more disordered because of stronger springs. CONCLUSION: a spring constant of approximately 2 N/m exerting 0.10 N and greater significantly lengthens intestinal segments and stimulates intestinal structural changes at POD7 and 14. This suggests a decreased force is capable of inducing spring-mediated distraction enterogenesis.

Intestinal adaptation following spring insertion into a roux limb in mice.

BACKGROUND/PURPOSE: Intraluminal springs have recently been shown to lengthen segments of intestine in a process known as distraction enterogenesis. We hypothesized that biocompatible springs could be used to lengthen defunctionalized murine small intestine and would lead to identifiable intestinal adaptations at the molecular level. METHODS: Age and weight matched C57BL/6 mice underwent surgical insertion of nitinol spring-loaded capsules into a Roux limb of jejunum. Segment lengths were measured at initial spring placement and at euthanasia after 14 and 21 days. Histology and gene expression of the Roux limb were evaluated at scarification and compared to untreated control segments. RESULTS: Intestinal segments loaded with compressed springs lengthened an average of 240%, which was significantly longer than control segments loaded with either empty capsules or uncompressed springs. Muscularis thickening was greater in spring-treated mice compared to controls without springs. Crypt depth and Lgr5+ expression was greater in mice that received compressed spring treatments when compared to control groups. CONCLUSIONS: Insertion of a compressed nitinol spring into a Roux limb results in significant intestinal lengthening, smooth muscle thickening, and Lgr5+ expression in a mouse model. The ability to increase small bowel length in a defunctionalized murine model may be used to understand the mechanism of distraction enterogenesis.

Mesenteric neovascularization during spring-mediated intestinal lengthening.

BACKGROUND: Short gut syndrome, a condition characterized by inadequate absorption of nutrients owing to decreased bowel length, has minimal avenues for treatment. We have proposed spring-mediated distraction enterogenesis to lengthen bowel in porcine jejunum as a treatment for short gut. We aim to evaluate the extent of mesenteric neovascularization in segments of lengthened bowel via spring-mediated enterogenesis. METHODS: Female juvenile Yucatan pigs underwent laparotomy and insertion of gelatin-encapsulated compressed nitinol springs, held in place with plication sutures, into the jejunum. At surgery and sacrifice, macroscopic mesenteric blood vessels were counted between the plication sites. Histologic samples of the mesentery were obtained to evaluate microscopic vasculature. RESULTS: A statistically significant increase in macroscopic mesenteric blood vessels was seen after intestinal lengthening (before: 1.9 ±â€¯0.7 vessels, after: 4.7 ±â€¯1.2 vessels, p = 0.001). A statistical significance is also seen in the density of arterioles (control: 3.0 ±â€¯3.0 vessels/mm, spring: 7.0 ±â€¯9.0 vessels/mm, p = 0.01) and venules (control: 4.0 ±â€¯3.0 vessels/mm, spring: 8.0 ±â€¯8.0 vessels/mm, p = 0.003). CONCLUSION: Intestinal segments lengthened by intraluminal springs demonstrated total greater number of macroscopic vessels and microscopic blood vessels per length of mesentery as compared to control. This suggests local changes within the mesentery to recruit blood supply to growing intestine. LEVEL OF EVIDENCE: N/A TYPE OF STUDY: Treatment study.

Human skin-derived precursor cells xenografted in aganglionic bowel.

PURPOSE: One in 5000 newborns is diagnosed with Hirschsprung disease each year in the United States. The potential of employing neural crest stem cells to restore the enteric nervous system has been investigated. Skin-derived precursor cells (SKPs) are multipotent progenitor cells that can differentiate into neurons and gliocytes in vitro and generate enteric ganglion-like structures in rodents. Here we examined the behavior of human SKPs (hSKPs) after their transplantation into a large animal model of colonic aganglionosis. METHODS: Juvenile minipigs underwent a chemical denervation of the colon to establish an aganglionosis model. The hSKPs were generated from human foreskin and were cultured in neuroglial-selective medium. Cells were labeled with a fluorescent dye and were injected into the porcine aganglionic colon. After one week, transplanted hSKPs were assessed by immunofluorescence for markers of multipotency and neuroglial differentiation. RESULTS: In culture, hSKPs expressed nestin and S100b indicative of neuroglial precursors. After xenografting in pigs, hSKPs were identified in the myenteric and submucosal plexuses of the colons. The hSKPs expressed nestin and early neuroglial differentiation markers. CONCLUSIONS: Human SKPs transplanted into aganglionic colon demonstrated immunophenotypes of neuroglial progenitors, suggesting their potential use for Hirschsprung disease.

Electroacupuncture to Increase Neuronal Stem Cell Growth.

Background: Neuropathic intestinal disorders continue to pose a significant burden, and current treatment options do not target the underlying cellular deficiencies. The goal of this study is to determine whether acupuncture and electroacupuncture (EA) can affect the growth of neuronal cells. Methods: Three groups of Lewis rats received 25 minutes of acupuncture twice a week for 10 weeks. The 3 groups of rats received treatment with either sham acupuncture (SA), real acupuncture (RA), or EA. After 10 weeks of treatment, skin and intestinal tissue were collected and analyzed for histology and mRNA expression of neuronal marker genes. Results: Compared with rats that received SA, rats that received RA and EA showed a significant increase in the mRNA expression levels of multiple neuronal genes in the skin. No significant histologic changes were seen. Conclusions: Acupuncture and EA result in significant changes in the expression of genes implicated as markers for neural stem cells, neural cell development, and neurons. This may, therefore, provide a novel avenue for developing treatments in patients suffering from intestinal aganglionic and neuropathic diseases.

Biomechanical signaling and collagen fiber reorientation during distraction enterogenesis.

Distraction enterogenesis has been extensively studied as a potential treatment for short bowel syndrome, which is the most common subset of intestinal failure. Spring distraction uses an intraluminal axial mechanical force to stimulate the growth and elongation of the small intestine. The tissue close to the distracted intestinal segment may also experience signaling to grow. In this study we examined the effects of distraction enterogenesis at different post-operative days on the thickness of small intestinal layers in the intestine proximal and distal to the distracted segment, as well as how the submucosal collagen fibers were reoriented. It was observed that not only different layers of intestine wall in distracted segment showed thickening due to the applied mechanical force but also adjacent tissues in both distal and proximal directions were impacted significantly where they showed thickening as well. The orientation of collagen fibers in submucosa layer was also significantly impacted due to the mechanical force in both distracted and adjacent tissue. The effect of the applied mechanical force on the main distracted tissue and the radial growth of the adjacent tissue strongly suggest actions of paracrine signaling.

Optimization of In-Continuity Spring-Mediated Intestinal Lengthening.

BACKGROUND: Spring-mediated intestinal lengthening has been studied in numerous animal models to effectively achieve up to a 3-fold increase in length. In this study we are interested in optimizing this method of spring lengthening. METHODS: Juvenile mini-Yucatan pigs underwent laparotomy for spring implantation. Springs were secured by plicating the intestine around the springs. In one set of experiments, varying degrees of plication were compared to determine the necessary narrowing needed to confine the spring. In another set of experiments, dissolvable sutures were used for the plication to allow for spontaneous spring passage postoperatively. Intestinal segments were retrieved and evaluated for lengthening and histological changes. RESULTS: Pigs tolerated their diet advancement to a regular diet postoperatively. 10% plication resulted in a 1.3-fold increase in length, while 50% plication resulted in a 2.7-fold increase in length (p<0.05). At two months postoperatively, the majority of springs had safely passed out of the intestine. All lengthened intestine showed significant growth histologically. CONCLUSIONS: A 50% reduction in lumen diameter achieves optimal spring-mediated intestinal lengthening. Springs can safely pass out of the intestine, thus avoiding a second operation for spring removal. These results may be important in developing future therapies for short bowel syndrome. LEVEL OF EVIDENCE: Level I experimental study.

Autologous Transplantation of Skin-Derived Precursor Cells in a Porcine Model.

BACKGROUND: Hirschprung's disease is characterized by aganglionic bowel and often requires surgical resection. Cell-based therapies have been investigated as potential alternatives to restore functioning neurons. Skin-derived precursor cells (SKPs) differentiate into neural and glial cells in vitro and generate ganglion-like structures in rodents. In this report, we aimed to translate this approach into a large animal model of aganglionosis using autologous transplantation of SKPs. METHODS: Juvenile pigs underwent skin procurement from the shoulder and simultaneous chemical denervation of an isolated colonic segment. Skin cells were cultured in neuroglial-selective medium and labeled with fluorescent dye for later identification. The cultured SKPs were then injected into the aganglionic segments of colon, and the specimens were retrieved within seven days after transplantation. SKPs in vitro and in vivo were assessed with histologic samples for various immunofluorescent markers of multipotency and differentiation. SKPs from the time of harvest were compared to those at the time of injection using PCR. RESULTS: Prior to transplantation, 72% of SKPs stained positive for nestin and S100b, markers of neural and glial precursor cells of neural crest origin, respectively. Markers of differentiated neurons and gliocytes, TUJ1 and GFAP, were detected in 47% of cultured SKPs. After transplantation, SKPs were identified in both myenteric and submucosal plexuses of the treated colon. Nestin co-expression was detected in the SKPs within the aganglionic colon in vivo. Injected SKPs appeared to migrate and express early neuroglial differentiation markers. CONCLUSIONS: Autologous SKPs implanted into aganglionic bowel demonstrated immunophenotypes of neuroglial progenitors. Our results suggest that autologous SKPs may be potentially useful for cell-based therapy for patients with enteric nervous system disorders. TYPE OF STUDY: Basic science.

Double plication for spring-mediated in-continuity intestinal lengthening in a porcine model.

BACKGROUND: Short bowel syndrome is a condition with substantial morbidity and mortality, yet definitive therapies are lacking. Distraction enterogenesis uses mechanical force to "grow" new intestine. In this study, we examined whether intestinal plication can be used to safely achieve spring-mediated intestinal lengthening in a functioning segment of jejunum in its native position. METHODS: A total of 12 juvenile, miniature Yucatan pigs underwent laparotomy to place either compressed springs or expanded springs within a segment of jejunum (n = 6 per group). The springs were secured within the jejunum by performing intestinal plication to narrow the intestinal lumen around the spring. After 3 weeks, the jejunum was retrieved and examined for lengthening and for histologic changes. RESULTS: There were no intraoperative or postoperative complications, and the pigs tolerated their diets and gained weight. Segments of jejunum containing expanded springs showed no significant change in length over the 3 weeks. In contrast, jejunum containing compressed springs showed nearly a 3-fold increase in length (P < .001). Histology of the retrieved jejunum showed a significant increase in thickness of the muscularis propria and in crypt depth relative to normal jejunum. CONCLUSION: Intestinal plication is effective in securing endoluminal springs to lengthen the jejunum. This approach is a clinically relevant model because it allows for normal GI function and growth of animals during intestinal lengthening, which may be useful in lengthening intestine in patients with short bowel syndrome.

Intestinal lengthening via multiple in-continuity springs.

BACKGROUND: Short bowel syndrome is a debilitating condition with few effective treatments. Spring-mediated distraction enterogenesis can be used to lengthen intestine. The purpose of this study is to determine whether multiple springs in series can safely increase the total amount of lengthening. METHODS: Juvenile mini-Yucatan pigs each received three nitinol springs placed within their jejunum. Plication was used to narrow the intestine around each spring to secure them. Compressed springs were used in the experimental group, while uncompressed springs were used in the control group. The intestine was examined 3 weeks later for lengthening and histologic changes. RESULTS: All pigs tolerated diets postoperatively with continued weight gain, and no dilation or obstruction of the intestine was observed. Segments of intestine that contained compressed springs had a significant increase in length from 2.5 cm to 3.9 ±â€¯0.2 cm per spring, compared to segments containing control springs that showed no change (p < 0.001). CONCLUSIONS: Intestinal plication can be safely used to secure multiple springs in series to achieve intestinal lengthening without compromising intestinal function. Using several springs at once allows for a greater amount of total lengthening. This is a promising model that has potential in the treatment of short bowel syndrome.